The object of this research is to understand the role of specific lesions in DNA effecting cell death. We have developed a radioimmunoassay capable of detecting photoproducts in the DNA of mammalian cells at biologically relevant doses. This tool will now be used in combination with monoclonal antibodies to UV-irradiated DNA in order to determine the importance of different UV photoproducts. Even though we have shown that the antiserum (Ab) recognizes primarily thymine dimers, loss of Ab-binding sites during DNA repair does not correlate with the loss of sensitivity to dimer specific endonucleases. We have hypothesized that loss of Ab-binding results from aglycosylation of the dimers. This will be tested by measuring the capacity of "repaired" DNA to act as a substrate for ADP endonuclease. In order to further our understanding of the initial steps in DNA repair, this assay will also be applied to DNA from UV-sensitive mutants from CHO cells, in which loss of Ab-binding sites does not occur. Since dimers are modified in such a way as to lose their sensitivity to existing Ab's, monoclonal Ab's will be made to "repaired" CHO cell DNA. The inability of damaged DNA to bind these Ab's may then be indicative of dimer excision. Similarly, the distribution of residual dimers within the genome is still unresolved. Using a physiologically-meaningful chromatin fractionation technique no difference in the rate of loss of Ab-binding sites in the different fractions was evident. Since it is probable that this assay does not measure complete removal of dimers, the development of Ab's to "modified" dimers may elucidate this point. Further work on the characterization of our chromatin fractions is also proposed and involves the use of cDNA from a specific gene product as a probe for transcribing DNA sequences.